Signaling protein abundance modulates the strength of the spindle assembly checkpoint.

During mitosis, unattached kinetochores in a dividing cell signal to the spindle assembly checkpoint (SAC) to delay anaphase onset and prevent chromosome missegregation.1,2,3,4 The signaling activity of these kinetochores and the likelihood of chromosome missegregation depend on the amount of SAC signaling proteins each kinetochore recruits.5,6,7,8 Therefore, factors that control SAC protein recruitment must be thoroughly understood. Phosphoregulation of kinetochore and SAC signaling proteins due to the concerted action of many kinases and phosphatases is a significant determinant of the SAC protein recruitment to signaling kinetochores.9 Whether the abundance of SAC proteins also influences the recruitment and signaling activity of human kinetochores has not been studied.8,10 Here, we reveal that the low cellular abundance of the SAC signaling protein Bub1 limits its own recruitment and that of BubR1 and restricts the SAC signaling activity of the kinetochore. Conversely, Bub1 overexpression results in higher recruitment of SAC proteins, producing longer delays in anaphase onset. We also find that the number of SAC proteins recruited by a signaling kinetochore is inversely correlated with the total number of signaling kinetochores in the cell. This correlation likely arises from the competition among the signaling kinetochores to recruit from a limited pool of signaling proteins, including Bub1. The inverse correlation may allow the dividing cell to prevent a large number of signaling kinetochores in early prophase from generating an overly large signal while enabling the last unaligned kinetochore in late prometaphase to signal at the maximum strength.

BubR1 recruitment to the kinetochore via Bub1 enhances spindle assembly checkpoint signaling.

During mitosis, unattached kinetochores in a dividing cell activate the spindle assembly checkpoint (SAC) and delay anaphase onset by generating the anaphase-inhibitory mitotic checkpoint complex (MCC). These kinetochores generate the MCC by recruiting its constituent proteins, including BubR1. In principle, BubR1 recruitment to signaling kinetochores should increase its local concentration and promote MCC formation. However, in human cells BubR1 is mainly thought to sensitize the SAC to silencing. Whether BubR1 localization to signaling kinetochores by itself enhances SAC signaling remains unknown. Therefore, we used ectopic SAC activation (eSAC) systems to isolate two molecules that recruit BubR1 to the kinetochore, the checkpoint protein Bub1 and the KI and MELT motifs in the kinetochore protein KNL1, and observed their contribution to eSAC signaling. Our quantitative analyses and mathematical modeling show that Bub1-mediated BubR1 recruitment to the human kinetochore promotes SAC signaling and highlight BubR1's dual role of strengthening the SAC directly and silencing it indirectly.

Stu2 acts as a microtubule destabilizer in metaphase budding yeast spindles.

The microtubule-associated protein Stu2 (XMAP215) has the remarkable ability to act either as a polymerase or as a destabilizer of the microtubule plus end. In budding yeast, it is required for the dynamicity of spindle microtubules and also for kinetochore force generation. To understand how Stu2 contributes to these distinct activities, we analyzed the contributions of its functional domains to its localization and function. We find that Stu2 colocalizes with kinetochores using its TOG domains, which bind GTP-tubulin, a coiled-coil homodimerization domain, and a domain that interacts with plus-end interacting proteins. Stu2 localization is also promoted by phosphorylation at a putative CDK1 phosphorylation site located within its microtubule-binding basic patch. Surprisingly, however, we find that kinetochore force generation is uncorrelated with the amount of kinetochore-colocalized Stu2. These and other data imply that Stu2 colocalizes with kinetochores by recognizing growing microtubule plus ends within yeast kinetochores. We propose that Stu2 destabilizes these plus ends to indirectly contribute to the "catch-bond" activity of the kinetochores.

Health at Every Size College Course Reduces Dieting Behaviors and Improves Intuitive Eating, Body Esteem, and Anti-Fat Attitudes.

OBJECTIVE: To investigate the effects of a Health at Every Size general education course on intuitive eating, body esteem (BES), cognitive behavioral dieting scores, and anti-fat attitudes of college students. METHODS: Quasi-experimental design with 149 students in intervention (45), comparison (66), or control (46) groups. Analysis of variance and post hoc Tukey adjusted tests were used. RESULTS: Mean scores for total general education course on intuitive eating (P < .001), unconditional permission to eat (P < .001), reliance on hunger (P < .001), cognitive behavioral dieting scores (P < .001), BES appearance (P = .006), BES weight (P < .001), and anti-fat attitudes (P < .001) significantly improved from pre to post in the intervention group compared with control and comparison groups. CONCLUSION AND IMPLICATIONS: Students in the Health at Every Size class improved intuitive eating, body esteem, and anti-fat attitudes and reduced dieting behaviors compared with students in the control and comparison groups.

Does a high-protein diet improve weight loss in overweight and obese children?

OBJECTIVE: To evaluate the effect of a high-protein diet on anthropometry, body composition, subjective appetite, and mood sensations in overweight and obese children attending a residential weight-loss camp. RESEARCH METHODS AND PROCEDURES: Children (120; BMI, 33.1 +/- 5.5 kg/m(2); age, 14.2 +/- 1.9 years) were randomly assigned to either a standard or high-protein diet group (15% vs. 22.5% protein, respectively). All children were assessed at baseline and at the end of the camp for anthropometry, body composition, blood pressure, biochemical variables (n = 27), and subjective appetite and mood sensations (n = 50). RESULTS: Attendance at the weight-loss camp resulted in significant improvements in most measures. Campers lost 5.5 +/- 2.9 kg in body weight (p < 0.001) and 3.8 +/- 5.4 kg in fat mass (p < 0.001) and reduced their BMI standard deviation score by 0.27 +/- 0.1 (p < 0.001) and their waist circumference by 6.6 +/- 2.8 cm (p < 0.001). Subjective sensations of hunger increased significantly over the camp duration, but no other changes in appetite or mood were observed. There were no significant differences between the two diets on any physical or subjective measures. DISCUSSION: Weight-loss camps are effective in assisting children to lose weight and improve on a range of health outcomes, independently of the protein content of the diet. The implications of an increase in hunger associated with weight loss needs to be considered. Further work is warranted to investigate whether higher levels of dietary protein are feasible or effective in longer-term weight-loss interventions of this type.